Growing Human Parathyroids in a Microphysiological System: A Novel Approach to Understanding and Developing New Treatments for Hyperparathyroidism.

We developed a novel model for studying hyperparathyroidism by growing ex vivo 3-dimensional human parathyroids as part of a microphysiological system (MPS) that mimics human physiology. The purpose of this study was to validate the parathyroid portion of the MPS. We prospectively collected parathyroid tissue from 46 patients with hyperparathyroidism for growth into pseudoglands. We evaluated pseudogland architecture and calcium responsiveness. Following 2 weeks in culture, dispersed cells successfully coalesced into pseudoglands ∼500-700 µm in diameter that mimicked the appearance of normal parathyroid glands. Functionally, they also appeared similar to intact parathyroids in terms of organization and calcium-sensing receptor expression. Immunohistochemical staining for calcium-sensing receptor revealed 240-450/cell units of mean fluorescence intensity within the pseudoglands. Finally, the pseudoglands showed varying levels of calcium responsiveness, indicated by changes in parathyroid hormone (PTH) levels. In summary, we successfully piloted the development of a novel MPS for studying the effects of hyperparathyroidism on human organ systems. We are currently evaluating the effect of PTH on adverse remodeling of tissue engineered cardiac, skeletal, and bone tissue within the MPS.

Water Clear Cell Adenoma of the Parathyroid Gland: A Forgotten Cause of Primary Hyperparathyroidism.

Water clear cell adenoma of the parathyroid gland is a rare neoplasm composed of large clear cells with foamy cytoplasm and mild nuclear pleomorphism, compressing the residual nonneoplastic parathyroid tissue. The differential diagnosis includes a variety of neoplasms with clear cell features. In this article, we provide an overview of the entity with a historical perspective, in order to help pathologists in distinguishing it from other neoplasms in their daily practice.

Is the ultimobranchial body a reality or myth: a study using serial sections of human embryos.

Reported morphologies of the ultimobranchial body had varied between researchers: a cluster of mitotic cells, a duct-like structure and a rosette-like cell mass. To clarify the true morphology, we studied tilted horizontal sections of 20 human embryos (crown-rump length 5-18 mm; 4-6 weeks). The sections displayed a ladder-like arrangement of the second to fourth endodermal pouches and, in 5 early embryos we found the fifth pouch attached to the fifth ectodermal groove near the fourth pharyngeal arch artery. The bilateral fifth pharyngeal pouches protruded anterolaterally to form a U-shaped lumen surrounding the arytenoid swelling. The third to fifth pouches were each characterized by a pedal-shaped inferior end. We identified several types of cell clusters as candidates for the ultimobranchial body, but morphologically most of them were, to various degrees, likely to correspond to the blind end of the lower pouch when cut tangentially. Because of the topographical relation to the common carotid artery, a cyst-like structure with a cell cluster seemed to be the most likely candidate of the ultimobranchial body (a common anlage of the thymus and parathyroid). However, we were not able to deny a possibility that a certain plane cutting the pouch end incidentally provided such a cyst-like structure in sections. At any stage, the ultimobranchial body might not appear as a definite structure that is discriminated from others with routine staining. A concept of the ultimobranchial body might be biased by comparative anatomy that shows the ultimobranchial gland in adult birds and reptiles.

Identification of parathyroid glands: anatomical study and surgical implications.

While performing thyroid surgery, the unintentional lesion of parathyroid glands and laryngeal nerves results in a profound alteration in patient's quality of life. To minimize thyroid surgery morbidity, the surgeon must have an in-depth knowledge of the thyroid gland morphology and its anatomical relations in the anterior compartment of the neck. This work intended to simulate total thyroidectomies using cadaver parts and isolate fragments that may correspond to parathyroid glands. The thyroid glands and "eventual" parathyroid glands were then submitted to histological study. Ninety-two cadaver parts were used for macroscopic dissection. A total of 242 fragments were isolated, 154 of which were confirmed through histological study to be parathyroid glands. In 36 cases, all "eventual" parathyroid glands isolated during dissection were confirmed through histological verification. In 40 cases, some glands were confirmed. In 16 cases, none of the "eventual" parathyroid glands was confirmed. The 92 thyroid glands isolated during dissection were also submitted to histological study. In 21 thyroid glands, 16 parathyroid glands were identified in the histological cuts: 8 sub-capsular, 8 extra-capsular, 6 intra-thyroidal. There was no statistical difference between the dimensions of the parathyroid glands. Parathyroid gland identification and preservation are sometimes a challenge during thyroid surgery, difficulty that has been demonstrated during dissection of cadaver parts.

Age-related changes of the ultrastrucure in the parathyroid gland of the golden hamster.

We qualitatively and quantitatively investigated the parathyroid glands of golden hamsters aged 6, 12, 18, 24 and 30 months. Percent area of rER in the parathyroid gland in golden hamsters at 24 months of age was significantly higher when compared to 6 and 12 months of age, and the percent area at 30 months of age was significantly higher when compared to 12 months of age, but there were no significant differences between 24 and 30 months of age. Percent area of the Golgi apparatus at 24 and 30 months of age was significantly higher when compared to 6, 12 and 18 months of age. Ultrastructurally, we believe that in the parathyroid gland of the golden hamster, synthesis and release of parathyroid hormone increase gradually from 6 to 24months of age and are maintained from 24 to 30 months of age.

Effect of maternal nicotine/thiocyanate exposure during gestational period upon pituitary, thyroid and parathyroid function/morphology of 1-month-old rat offspring.

INTRODUCTION: Impact of in utero exposure to nicotine, on the structure of the thyroid-pituitary axis and the parathyroid glands have been examined in 1-month-old rats and compared with that of thiocyanate. MATERIALS AND METHODS: Three pregnant female groups were used; control, nicotine and thiocyanate. Treatment started from gestation day (4-20) and the specimens were harvested from the male offspring of all groups at the age of 1 month and processed for light, electronmicroscopic and immunohistochemical examination. Total triiodothyronine (tT3), total thyroxine (tT4) and total thyrotropin (TSH) were quantitatively determined in serum. RESULTS: Both nicotine and thiocyanate activated the thyroid follicular cells, with an increase in height (about 30 %) and a negative feedback on the pituitary thyrotrophs which revealed a reduction in the number of cytoplasmic secretory granules, particularly the thiocyanate group. However, in thiocyanate group there was signs of impaired secretory activity of the thyroid gland. The arbitrary area of parathyroid chief cells, increased (about 45 %) particularly in nicotine group, with signs of reduced activity and a positive feedback on the parafollicular cells which revealed hypertrophy, proliferation (25 %) and increased intensity of positive immunohistochemical reaction for calcitonin. CONCLUSION: Nicotine impaired chief parathyroid cells activity and consequently activated parafollicular cells. Thiocyanate reduced pituitary thyrotrophs activity, whereas both nicotine and thiocyanate increased thyroid follicular cells activity. This impact of in utero exposure persisted for 1-month postnatal.

Genistein affects parathyroid gland and NaPi 2a cotransporter in an animal model of the andropause.

This study aimed to examine the effects of genistein on the structural and functional changes in parathyroid glands (PTG) and sodium phosphate cotransporter 2a (NaPi 2a) in orchidectomized rats. Sixteen-month-old Wistar rats were divided into sham-operated (SO), orchidectomized (Orx) and genistein-treated orchidectomized (Orx+G) groups. Genistein (30 mg/kg/day) was administered subcutaneously for 3 weeks, while the controls received vehicle alone. PTG was analyzed histomorphometrically, while the expressions of NaPi 2a mRNA/protein levels from kidneys were determined by real time PCR and Western blots. Serum and urine parameters were determined biochemically. The PTG volume in Orx rats was increased by 30% (p<0.05), compared to the SO group. Orx+G treatment increased the PTG volume by 35% and 75% (p<0.05) respectively, comparing to Orx and SO animals. Orchidectomy led to increment of serum PTH by 27% (p<0.05) compared to the SO group, Orx+G decreased it by 18% (p<0.05) comparing to Orx animals. NaPi 2a expression in Orx animals was reduced in regards to its abundance in SO animals, although it was increased in Orx+G group compared to the Orx. Phosphorus urine content of Orx animals was raised by 12% (p<0.05) compared to that for the SO group, while Orx+G induced a 17% reduction (p<0.05) in regards to Orx animals. Our study shows that Orx increases PTG volume and serum PTH level, while protein expression of NaPi 2a is reduced. Application of genistein attenuates the orchidectomy-induced changes in serum PTH level, stimulates the expression of NaPi 2a and reduces urinary Pi excretion, implying potential beneficial effects on andropausal symptoms.

Effects of time on ultrastructural integrity of parathyroid tissue before cryopreservation.

BACKGROUND: Cryopreservation of parathyroid tissue is used in the surgical treatment of secondary hyperparathyroidism. After surgical resection, the tissue is temporarily maintained in a cell culture solution until it arrives at the specialized laboratory where the cryopreservation process will take place. The present study evaluates the time that the human hyperplastic parathyroid gland tissue can wait before cryopreservation, based on parathyroid cell ultrastructural integrity. METHODS: This prospective study included 11 patients who underwent total parathyroidectomy with heterotopic autotransplantation and cryopreservation of parathyroid tissue fragments. Part of the tissue was kept in cell culture solution at 4 °C. Five time periods between 2 and 24 h were defined, and parathyroid fragments were kept in the solution for that length of time. At the end of each period, the fragments were removed from the transport solution, fixed, and prepared for ultrathin sections. RESULTS: Of the 11 cases studied, 10 showed ultrastructural findings consistent with cellular viability in tissue fragments that remained in the transport solution up to 12 h. Electron microscopy revealed that cell adhesion and the integrity of plasma membranes, nuclei, and mitochondria were preserved in one case for up to 24 h. Changes in mitochondrial structure represented the most constant ultrastructural damage seen in the cases studied, in addition to the presence of edema and cell vacuoles. CONCLUSIONS: Analysis of the ultrastructure of hyperplastic parathyroid gland tissue showed that ultrastructural integrity was in most cases properly maintained in fragments stored up to 12 h in a cell culture solution at 4 °C.

Development and prevention of morphologic and ultrastructural changes in uremia-induced hyperplastic parathyroid gland.

The detailed ultrastructural changes of uremia-induced hyperplastic parathyroid gland and the effects of current medical treatments for secondary hyperparathyroidism were investigated. Marked enlargement of parathyroid cell with accumulation of mitochondria and lipids and a significant increase in the thickness of the pericapillary area with increased fibrosis and appearance of fibroblast like cells were noted in the hyperplastic gland caused by uremia and phosphate retention. These ultrastructural changes and biochemical findings indicating hyperparathyroidism were significantly suppressed by all of the treatment using phosphate restriction, calcitriol, and cinacalcet. The characteristic ultrastructural changes, including the morphologic evidence of nodule formation, were indicated.

Stimulating parathyroid cell proliferation and PTH release with phosphate in organ cultures obtained from patients with primary and secondary hyperparathyroidism for a prolonged period.

The pathogenesis of primary hyperparathyroidism (I degrees -HPT) and secondary hyperparathyroidism (II degrees -HPT) remains to be elucidated. To characterize their pathophysiology, we investigated the effects of calcium and phosphate on cell proliferation and PTH release in an organ culture of parathyroid tissues. Dissected parathyroid tissues obtained from patients with I degrees -HPT (adenoma) or II degrees -HPT (nodular hyperplasia) were precultured on a collagen-coated membrane for 1-4 week. After changing the medium for one containing various concentrations of phosphate, PTH release and [(3)H]thymidine incorporation were studied. In contrast to dispersed parathyroid cells cultured in a monolayer, calcium decreased PTH release in a concentration-dependent manner in parathyroid tissues. Furthermore, when parathyroid tissues obtained from II degrees -HPT were precultured for 1-4 weeks, PTH release and parathyroid cell proliferation were significantly increased in high-phosphate medium. These phosphate effects were also observed to a lesser extent in parathyroid tissues obtained from I degrees -HPT, but there was no significant difference between I degrees -HPT and II degrees -HPT. Microarray analyses revealed that mRNA levels of PTH, CaSR, and VDR were well preserved, and several growth factors (e.g. TGF-beta1-induced protein) were abundantly expressed in II degrees -HPT. Using organ cultures of hyperparathyroid tissues, in which PTH release and CaSR are well preserved for a prolonged period, we have demonstrated that phosphate stimulates parathyroid cell proliferation not only in II degrees -HPT but also in I degrees -HPT. Although the mechanism responsible for phosphate-induced cell proliferation remains to be elucidated, our in vitro findings suggest that both parathyroid tissues preserve to some extent a physiological response system to hyperphosphatemia as observed in normal parathyroid cells.

Responses of parathyroid gland, C cells, and plasma calcium and inorganic phosphate levels in rat to sub-lethal heroin administration.

In order to record the effects of heroin on plasma calcium (Ca) and inorganic phosphate (Pi) levels as well as parathyroid gland and C cells, two sub-lethal doses (0.50 LD50 and 0.75 LD50) of the drug were administered intramuscularly in Rattus norvegicus for 30 days. Plasma Ca level of control rats ranged between 9.53 +/- 0.32 - 9.88 +/- 0.22 mg 100 ml(-1) while plasma Pi concentration fluctuated between 4.55 +/- 0.18 - 4.71 +/- 0.24 mg 100 ml(-1). Sub-lethal heroin administration induced progressive increase in plasma Ca level during the first seven days (p < 0.001), thereafter the level declined on day 15 and 30. However plasma Pi level of the heroin-treated rats registered increase with the peak value (p < 0.001) on day 30. The treatment elicited degenerative changes in parathyroid gland as evident by cytoplamic vacuolization, presence of more pycnotic nuclei and occurrence of patchy areas among the chief cells. Degenerative changes were also noticed in cristae of mitochondria, Golgi complex and endoplasmic reticulum. There was decrease in chromatin material in the nucleus and loss of hormone granules in the cytoplasm. Oxyphil cells of the heroin-treated rat depicted dilation of endoplasmic reticulum and damaged cristae. Sub-lethal heroin administration in the rat for 30 days induced dilation in endoplasmic reticulum and loss of secretory granules in C cells.

Responses of serum calcium and inorganic phosphate levels as well as parathyroid gland and calcitonin producing C cells of Rattus norvegicus to Mipcin administration.

Serum calcium (Ca) level of Rattus norvegicus ranged between 13.08 +/- 0.41 - 13.25 +/- 0.39 mg/100 ml whereas serum inorganic phosphate (Pi) concentration varied between 4.21 +/- 0.28 - 4.33 +/- 0.26 mg/100 ml. Sublethal (0.50 LD50 and 0.75 LD50) administration of Mipcin induced a progressive dose-dependent decline in serum Ca level in the rat which was statistically significant at 7 and 14 days. Serum inorganic phosphate level of the treated rats did not exhibit significant fluctuation during the entire course of investigation. Parathyroid chief cells of the experimental rats exhibited degranulation, vacuolation, loss of secretory (hormone) granules and lipid droplets, decreased chromatin in nuclei and damages in the endoplasmic reticulum as well as cristae of mitochondria at 14 days of the treatment. Not much of changes could be seen in the oxyphil cells of parathyroid as well as thyroid C cells of the Mipcin-treated rats.

Heterotopic parathyroid inclusion in a cervical lymph node.

BACKGROUND: During the pathologic examination of neck dissection specimens, unexpected findings within the lymph nodes may be discovered. Such findings may include benign epithelial inclusions, a second primary tumor, or chronic inflammatory diseases. METHODS: We report a case of a 59-year-old man who underwent a laryngectomy and bilateral neck dissection for a right transglottic squamous carcinoma of the larynx. During the procedure, a large lymph node measuring 2.5 x 2.2 x 0.8 cm was found at the right level IV. RESULTS: Histologic examination of the neck dissection specimen revealed benign parathyroid inclusions in the enlarged lymph node. The heterotopic cells expressed parathyroid hormone. This case represents a unique example of heterotopic parathyroid inclusions in a cervical lymph node. CONCLUSION: Parathyroid tissue should be included in the differential diagnosis of cervical intranodal epithelial inclusions.

Seasonal changes in plasma calcium and inorganic phosphate levels in relation to parathyroid structure of the grey quail, Coturnix coturnix coturnix Linnaeus.

Plasma Ca concentration (annual mean) in males Coturnix cotumix coturnix was 10.27 +/- 0.14 mg/100 ml while it was slightly higher (11.85 +/- 0.15 mg/100 ml) among females. Plasma Pi levels (annual mean) in males and females were 5.62 +/- 0.12 mg/100 ml and 6.52 +/- 0.20 mg/100 ml, respectively. While the males did not exhibit marked fluctuation in plasma Ca and Pi levels either in winter or summer, the females did record significant elevation in the levels of both these electrolytes during breeding season. The peak values of plasma Ca (17.66 +/- 0.38 mg/100 ml) and Pi (8.64 +/- 0.22 mg/100 ml) in females were observed during June. Parathyroid gland of the grey quail exhibited hyperactivity (hypertrophy and hyperplasia) during breeding season, however, the activity was more conspicuous among females than in males. The maximum increase in cell and nuclear diameters were observed in females during May-July. The follicles were also filled with AF- and PAS-positive materials during these months. The glands depicted signs of hypoactivity and degeneration during peak winter season (November-December) as evident by decrease in cell and nuclear diameters as well as vacuolation in the chief cells.

Parathyroid lipoadenomas: a rare cause of primary hyperparathyroidism.

OBJECTIVE: To review one institution's experience with parathyroid lipoadenomas and to report the associated clinical characteristics. METHODS: We present a case series of parathyroid lipoadenomas. A parathyroid lipoadenoma was defined as a single adenoma with more than 50% fat on histologic examination in conjunction with primary hyperparathyroidism and resolution of hypercalcemia postoperatively. Patients who fulfilled the diagnostic criteria were identified from the surgical pathology files of the Mayo Clinic. RESULTS: Five cases of parathyroid lipoadenomas, including 1 oxyphil lipoadenoma, were identified during the period from 1971 to 2001. The clinical picture of the study subjects resembled that of a typical patient with primary hyperparathyroidism. Parathyroid lipoadenomas were identified in 3 women (60%) and 2 men (40%), and only 1 patient presented with possible hypercalcemia-related symptoms of nephrolithiasis and hip fracture, leading to diagnosis. The mean serum calcium concentration was 11.1 mg/dL. Preoperatively, all study subjects had elevation of serum parathyroid hormone levels. Two of 3 patients (67%) had the tumor identified preoperatively by neck ultrasonography. CONCLUSION: A parathyroid lipoadenoma is a rare cause of primary hyperparathyroidism. The clinical features of this pathologic entity are similar to those of the more common pathologic variants of parathyroid disease associated with primary hyperparathyroidism.

Ultrastructure of the water-clear cell in the parathyroid gland of SAMP6 mice.

Although the parathyroid water-clear cell is very rare, it has clinical significance because of its association with parathyroid hyperplasia or adenoma. SAMP6, a substrain of senescence-accelerated mouse, was developed as an animal model for senile osteoporosis. We investigated the morphology of the parathyroid glands in SAMP6 and age-matched normal mouse SAMR1. The parathyroid water-clear cells, which contained numerous vacuoles and the crystalloid inclusions, were found in SAMP6 mice at 5, 8 and 12 months of age. It was noted that the number of water-clear cells increased with aging, which are fairly consistent with the change of the serum parathyroid hormone (PTH) level. We did not find any water-clear cells in the parathyroid glands of SAMR1 mice. The existence of water-clear cells may represent hyperfunction of the parathyroid glands in SAMP6.

Application trials of in situ hybridization at the electron microscopic level.

In situ hybridization (ISH) is a morphology-oriented technique for demonstrating the presence of specific nucleic acid sequences at chromosomal, cytological and histological levels. It is, however, sometimes difficult to recognize specific cell identity, early phase mRNA expression and alternative splicing because of the limited resolution of the light microscope. To overcome this limitation, we developed an improved technique for ISH at the electron microscopic level, in which pre-embedding hybridization with a non-radioactively labeled probe was used, followed by post-embedding immunoglobulin gold colloid staining. By applying this technique, early phase bone morphogenetic protein-3 mRNA in the nuclei and cytoplasm was successfully demonstrated in a differentiating chondrocytic cell lineage. Moreover, with oligo-DNA probes specific for alternative spliced forms of parathyroid hormone-related protein mRNA, we demonstrated such forms in a hyperplastic parathyroid gland attributed to renal failure.

Effects of diazinon on serum calcium and inorganic phosphate levels as well as ultrastructures of parathyroid and calcitonin cells of Rattus norvegicus.

Daily intramuscular administration of two doses--0.50 LD550 (150 mg/kg b w) and 0.75 LD50 (225 mg/kg b w) of diazinon for 14 days induced progressive hypocalcemia without altering serum inorganic phosphate value in Rattus norvegicus. Hypocalcemic response of the pesticide was dose- as well as time-dependent. Parathyroid chief cells of the experimental rats exhibited degranulation, vacuolation, loss of secretory granules and lipid droplets, decreased chromatin, and degenerative changes in endoplasmic reticulum, Golgi complex as well as cristae of the mitochondria. Not much of ultrastructural changes could be seen in the C cells of the rats treated with diazinon.

Ultrastructural and morphometrical evaluation of the parathyroid gland in iron-lactate-overloaded rats.

Iron lactate was given to Sprague-Dawley rats intravenously at the dosage of 10 mg/kg/day and the early effects on the parathyroid gland were examined ultrastructurally along with the blood level of parathyroid hormone (PTH) after single, 3-day or 6-day administration. Blood levels of electrolytes and other parameters related to osteoclast dynamics were also measured by blood chemistry and histopathology. The plasma parathyroid hormone (PTH) level was elevated in the single and 3-day dosing group but was reduced in the 6-day dosing group. Histopathologically, an increase of osteoclasts in the primary spongiosa was observed in the 3- and 6-day dosing groups. Image analysis of the parathyroid gland revealed that the average area of the storage granule decreased during a experimental period, with the number of storage granules decreasing in the 3- and 6-day dosing group. The chief cells of the parathyroid gland were moderately atrophied in the 6-day dosing group. These results demonstrate that iron lactate immediately promotes discharge of PTH from the storage granules after the treatment and induces an increase of osteoclasts in the primary spongiosa. The findings collectively suggest a pathophysiological mechanism of iron lactate-induced osteopenia in rats.

Nerve fibres in the parathyroid gland of the golden hamster (Mesocricetus auratus): immunohistochemical and ultrastructural investigations.

We investigated the morphology and the distribution of the nerve fibres in golden hamster (Mesocricetus auratus) parathyroid glands using antibodies to calcitonin gene-related peptide (CGRP) and substance P, and electron microscopy. CGRP-immunoreactive nerve fibres were densely distributed in the interstitial tissues and the capsules of the hamster parathyroid glands. Some nerve fibres were detected in close proximity of the parathyroid chief cells. The distribution pattern for substance P-immunoreactive nerve fibres was roughly the same as for CGRP-immunoreactive fibres. Ultrastructurally, we found numerous nerve fibres joining the blood vessels. Axon bundles were located adjacent to the smooth muscle cells of the arterioles. The axons formed structurally specialized neuromuscular junctions with the vascular smooth muscle cells. Some axons were in close vicinity to the parathyroid chief cells. These findings indicate that the hamster parathyroid gland contain CGRP and substance P, which may regulate the blood flow and the secretory activity of the gland.